摘要
为了探讨不同饲养层细胞对兔原始生殖细胞体外培养与传代的影响,从14-18 d的兔胎儿生殖嵴中分离得到原始生殖细胞（PGCs）,分别培养在小鼠胎儿成纤维细胞（MEF）、兔胎儿成纤维细胞（REF）饲养层上或与同源兔胎儿生殖脊成纤维细胞（HEFgr）、兔睾丸支持细胞（RSC）共培养。观察兔类EG集落的生长状态,并检测其碱性磷酸酶活性。结果表明：4种方法均能分离克隆出兔类EG集落,碱性磷酸酶染色均呈阳性,但是采用共培养的模式分离得到的集落明显多于传统的饲养层模式。与RSCs共培养得到的EG集落数最多,保持未分化的时间最长,并传了8代,与HEFgr共培养得到的EG集落数与RSCs相比差异不明显,但最高只传了6代。培养在MEF上的兔类EG也能较好的保持未分化状态,传了4代。培养在REF上的兔类EG集落数目较少,出现分化时间较早,只传了3代。因此可以用与RSCs或HEFgr共培养的方法传代培养兔PGCs。
To compare the effect of different feeder layers on subculture of rabbit primordial germ cells.Primordial germ cells（PGC） were isolated from 14～18 d rabbit embryonic genital ridges.And let the PGCs culture on mouse embryonic fibroblasts（MEF）,rabbit embryonic fibroblasts（REF） and co-cultured with the homogenous embryonic fibroblast from genital ridges（HEFgr）or co-cultured with Rabbit Sertoli cells（RSCs）.The rabbit EG like cell lines were observed and alkaline phosphatase（AKP） activity was detected respectively.The result is that EG cells were all able to gain in the four ways and all of them had the AKP activity,the manner of co-cultured was better than cultured depended on mitotically inactive feed layer.PGCs co-cultured with RSCs gained the more EG than other ways.They remained undifferentiated state for the longest period and cultured for 8 passages.There was not significant difference in PGCs co-cultured with the homogenous embryonic fibroblast from genital ridges and co-cultured with Rabbit Sertoli cells,but PGCs were just cultured for 6 passages.PGCs culture on mouse embryonic fibroblasts can also remained undifferentiated and cultured for 4 passages.PGCs culture on REF gained fewer EGs,they remained undifferentiated state were shorter and just cultured for 3 passages.So the manner of co-cultured with RSCs and co-cultured with the HEFgr can be used to isolation and cultivation rabbit PGCs.
出处
《西北农业学报》
CSCD
北大核心
2008年第3期37-41,共5页
Acta Agriculturae Boreali-occidentalis Sinica
基金
河南省杰出人才创新基金项目(0421002100)
作者简介
李义书（1982-），男，海南万宁人，在读硕士，主要从事动物胚胎工程研究。E-mail：liyishu2819@yahoo．com．cm．
通讯作者：王新庄，研究员，主要从事动物胚胎工程与发育生物学研究。E-mail：wangxinzhuang@yahoo．com．cn
